Project Summary/Abstract Current understanding of the diversity and geographic distribution of tick-borne pathogens in the United States remains limited. The majority of studies reported to date have focused on Borrelia burgdorferi sensu stricto, the causative agent of Lyme disease and the most commonly diagnosed vector-borne infection in the country. However, ticks in the United States also transmit several other pathogens that present additional, significant public health concerns, including spotted fever group Rickettsia spp., Ehrlichia spp., Babesia spp., and Anaplasma phagocytophilum. Moreover, novel viral and bacterial disease agents continue to be described from ticks. Here, we propose a small, self-contained project to perform a secondary analysis of an existing dataset, namely, several thousand ticks collected from more than one thousand dogs and several hundred cats in 49 of the 50 states in 2018 and 2019. Targeted, well- characterized assays will be performed to detect established tick-borne pathogens in the ticks. In addition, we will use next generation sequencing to characterize the metagenome of a subset of the ticks and then query the raw data with e-probes to discover novel agents that may be present. The findings from this work will inform future field efforts to more fully understand the geographic and temporal diversity of tick-borne pathogens in North America; greater understanding of the true public health menace presented by tick-borne infections is expected to result.